RESUMO
In the present study, we characterized a model system in which we examined the effects of human surfactant protein A (SP-A) on the uptake of a common human pulmonary pathogen, Pseudomonas aeruginosa, by a human monocytic/macrophage cell line, THP-1 cells. We found that SP-A significantly increases uptake of the bacteria in a dose-dependent manner. Bacterial uptake was temperature-dependent, because an effect of SP-A on bacterial uptake was observed at 37 degrees C and not at 4 degrees C. The continued presence of SP-A during the period when the bacteria and THP-1 cells were co-incubated was necessary for enhanced uptake. Pre-incubation of the bacteria or THP-1 cells with SP-A, followed by washing, abolished the effect of SP-A on bacterial uptake. The effect of SP-A could be inhibited by high concentrations of mannose, but was not affected by the removal or addition of lipopolysaccharide (LPS). Finally, we observed that the SP-A-mediated increase in uptake of P. aeruginosa by THP-1 cells was optimal in a narrow (100 mM and 150 mM) range of NaCl concentrations. We conclude that SP-A enhances the THP-1 cell-mediated uptake of P. aeruginosa in a manner dependent on temperature, the concentration of SP-A, and the concentration of NaCl.
Assuntos
Monócitos/efeitos dos fármacos , Fagocitose/efeitos dos fármacos , Proteolipídeos/farmacologia , Pseudomonas aeruginosa , Surfactantes Pulmonares/farmacologia , Cloreto de Sódio/farmacologia , Cápsulas Bacterianas/metabolismo , Fibrose Cística/complicações , Fibrose Cística/imunologia , Fibrose Cística/metabolismo , Relação Dose-Resposta a Droga , Glicosaminoglicanos/metabolismo , Humanos , Lipopolissacarídeos/farmacologia , Manitol/farmacologia , Manose/farmacologia , Monócitos/fisiologia , Concentração Osmolar , Fagocitose/fisiologia , Pneumonia Bacteriana/etiologia , Pneumonia Bacteriana/imunologia , Pneumonia Bacteriana/metabolismo , Infecções por Pseudomonas/etiologia , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/metabolismo , Proteína A Associada a Surfactante Pulmonar , Proteínas Associadas a Surfactantes Pulmonares , Solução Salina Hipertônica/farmacologia , TemperaturaRESUMO
In our study, surfactant protein (SP)-A was characterized in adult human trachea and bronchi. SP-A mRNA and protein were localized to serous cells in submucosal gland by in situ hybridization and immunohistochemistry, respectively. A 2.2 kb SP-A mRNA transcript was detected in tracheal tissues by Northern blot analysis. Primer extension analysis and gene-specific reverse transcriptase polymerase chain reaction (RT-PCR) revealed the predominance of SP-A2 mRNA. However, using nested PCR, we also detected low amounts of SP-A1 mRNA in the tracheal tissues. A approximately 35 kDa SP-A immunoreactive protein was detected in the tracheal tissues by immunoblot analysis and was shown to be modified by the addition of N-linked oligosaccharides. We conclude that submucosal glands in the conducting airways produce a novel SP-A protein with a molecular weight and post-translational modification similar to the SP-A produced in the distal lung. We speculate that this SP-A2 protein, like other serous secretions from airway submucosal glands, functions in local antimicrobial host defense mechanisms in the conducting airways.
Assuntos
Brônquios/metabolismo , Proteolipídeos/biossíntese , Surfactantes Pulmonares/biossíntese , Traqueia/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Northern Blotting , Feminino , Glicosilação , Humanos , Imuno-Histoquímica , Hibridização In Situ , Masculino , Pessoa de Meia-Idade , Mucosa/metabolismo , Reação em Cadeia da Polimerase , Proteína A Associada a Surfactante Pulmonar , Proteínas Associadas a Surfactantes Pulmonares , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Membrana Serosa/metabolismoRESUMO
Surfactant protein A (SP-A) is the major protein component of pulmonary surfactant, a material secreted by the alveolar type II cell that reduces surface tension at the alveolar air-liquid interface. The function of SP-A in the alveolus is to facilitate the surface tension-lowering properties of surfactant phospholipids, regulate surfactant phospholipid synthesis, secretion, and recycling, and counteract the inhibitory effects of plasma proteins released during lung injury on surfactant function. It has also been shown that SP-A modulates host response to microbes and particulates at the level of the alveolus. More recently, several investigators have reported that pulmonary surfactant phospholipids and SP-A are present in nonalveolar pulmonary sites as well as in other organs of the body. We describe the structure and possible functions of alveolar SP-A as well as the sites of extra-alveolar SP-A expression and the possible functions of SP-A in these sites.
Assuntos
Proteolipídeos/metabolismo , Alvéolos Pulmonares/metabolismo , Surfactantes Pulmonares/metabolismo , Animais , Proteínas Sanguíneas/antagonistas & inibidores , Proteínas Sanguíneas/farmacologia , Sistema Digestório/metabolismo , Orelha Média/metabolismo , Humanos , Lipopolissacarídeos/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Masculino , Proteínas Opsonizantes/metabolismo , Próstata/metabolismo , Proteolipídeos/antagonistas & inibidores , Proteolipídeos/genética , Proteolipídeos/farmacologia , Alvéolos Pulmonares/lesões , Alvéolos Pulmonares/microbiologia , Alvéolos Pulmonares/patologia , Proteína A Associada a Surfactante Pulmonar , Proteínas Associadas a Surfactantes Pulmonares , Surfactantes Pulmonares/antagonistas & inibidores , Surfactantes Pulmonares/genética , Surfactantes Pulmonares/farmacologia , Receptores de Superfície Celular/metabolismo , Baço/metabolismo , Tensão Superficial , Membrana Sinovial/metabolismo , Timo/metabolismoRESUMO
In the present study, pulmonary surfactant protein A (SP-A) messenger RNA (mRNA) and protein were characterized in adult rabbit middle ear and maxillary sinus. Fifteen adult rabbits were used for the study: 6 with evidence of acute middle ear infections and maxillary sinusitis, 6 with infections that were successfully treated with tetracycline, and 3 that were pathogen-free. We detected SP-A mRNA in maxillary sinus and middle ear tissues by Northern blot analysis and reverse transcriptase-polymerase chain reaction (RT-PCR). The RT-PCR also revealed the presence of SP-B and SP-C mRNA in middle ear and sinus tissues. We detected SP-A protein, of molecular weight approximately 29 and 70 kd, in middle ear and sinus tissues by immunoblot analysis. Unlike the SP-A protein present in the lung, the molecular weight of the SP-A protein present in the middle ear and paranasal sinus was not altered by digestion with an enzyme that cleaves N-linked carbohydrates. Immunostaining and in situ hybridization showed that SP-A protein and mRNA, respectively, were present in surface epithelial cells of the middle ear and in epithelial cells of submucosal glands in sinus tissues. These data provide the first evidence of the presence of pulmonary surfactant proteins in the paranasal sinuses and confirm previous reports of SP-A in the middle ear epithelium.